110 - Interrupting pro-inflammatory pulmonary-vascular cell interactions to reduce capillary leak in bronchiolitis

Poster Number: 110
Publication Number: 110.2730

Background: Bronchiolitis is a common cause of pediatric acute respiratory distress syndrome (PARDS). Microvascular endothelial dysfunction leads to pulmonary edema with hallmark symptoms of severe disease: difficulty breathing, hypoxemia from alveolar collapse, and gas diffusion reduction. Treatment for bronchiolitis induced PARDS is limited to intensive supportive care, as the pathogenesis of pulmonary microvascular dysfunction in bronchiolitis remains inadequately understood.

Objective: Our central hypothesis is that cells within the lung mitigate extra-alveolar Interleukin-1 (IL-1) signaling through endogenous inhibitors, reducing capillary leak and clinical hypoxemic respiratory failure. Our objective is to demonstrate that samples from the lungs of children with PARDS from bronchiolitis induce paracellular leak in cultured human pulmonary microvascular endothelial cells (HPMEC) which may be inhibited with IL-1 blockade.

Design/Methods: Tracheobronchial lavage (tBAL) samples were collected from 3 children with bronchiolitis induced PARDS on intubation and extubation. We performed single cell RNA-sequencing (scRNAseq), SomaScan proteomics and vascular function assays on all samples and analyzed with patient pairwise comparisons. Functional assays evaluated the effect of tBAL samples on HPMEC barrier function using electrical cell substrate impedance testing (ECIS). Gene expression analysis and immunofluorescence microscopy assessed cytoskeletal and junctional proteins of HPMEC treated with patient samples and IL-1 inhibitors.

Results: scRNAseq showed upregulated IL-1 signaling pathways in bronchial and alveolar cell types at intubation. Proteomic analysis revealed increased endogenous IL-1 inhibitors IL-1RN and IL-1R2 on extubation. HPMEC treatment with samples from intubation resulted in a sustained reduction in transendothelial electrical resistance (TEER) by 30%, disruption of tight junction proteins, and gap formation, while extubation samples slightly increased TEER. Pretreatment with IL-1 inhibitors IL-1RN and IL-1R2 reduced actin stress fibers. Elevated transcript levels of E-selectin and IL-6 indicated endothelial cell activation signifying a proinflammatory process.

Conclusion(s): Tracheobronchial fluid affects pulmonary endothelial cells through various vasoactive mediators. The observed functional and structural changes suggest that the mechanism underlying capillary leak is paracellular. Endothelial dysfunction plays an important role in the pathogenesis of pulmonary leak associated with acute lung injury in children. Identifying potential therapeutic targets is a vital aspect of future research.